Repetitive electroacupuncture causes prolonged increased met-enkephalin expression in the rVLM of conscious rats

Abstract

Enkephalinergic neurons in the rostral ventrolateral medulla (rVLM), an important presympathetic region in the brainstem, are activated by 30 min of low frequency (2 Hz) electroacupuncture (EA) at acupoints P5-P6, which overlie the median nerves. To more closely model the clinical application of acupuncture, we administered EA for 30 min twice over a 72 h period to unsedated conscious rats to examine its prolonged action. We hypothesized that repetitive EA would increase preproenkephalin mRNA and met-enkephalin in the rVLM of unsedated conscious rats. Rats received either EA (1-4 mA, 0.5 ms, 2 Hz) or sham stimulation (needle placement without electrical stimulation) twice at P5-P6 acupoints bilaterally. Preproenkephalin mRNA and its peptide met-enkephalin in the rVLM were measured 24 or 48 h after the final EA or sham procedure. Relative ratios of preproenkephalin mRNA levels (normalized with the 18S housekeeping gene) were almost doubled at 24h compared to sham (6.1 ± 0.79 vs. 3.1 ± 0.47). Met-enkephalin measured in rVLM tissue pooled from several rats exposed to the same treatment was increased by repeated EA by 68% after 24h and 51% after 48h, relative to sham. These findings suggest that repeated application of EA in the conscious rats enhances transcription and translation of enkephalin in rVLM for days. Since opioids in the rVLM contribute importantly to the action of EA on sympathetic outflow, this mechanism may contribute to the prolonged action of acupuncture on elevated blood pressure in patients.

Fig. 1

Site of punch biopsy taken from rVLM region for measurement of preproenkephalin mRNA and met-enkephalin. Left: a tissue section shows the site of punch biopsy (arrow) taken from rVLM region (12.24 mm caudal to Bregma). Right: the same rostral level of brain section indicated in the rats atlas (Paxinos and Watson, 2009). PY: Pyramid tract. Sp5: Spinal trigeminal tract. Sp5i: Spinal trigeminal nucleus, interpolar.

Fig. 2

Blood pressure and heart rate of a conscious rat before, during and after EA. The left and right arrows show the start and end times of EA.

Fig. 3

Relative preproenkephalin mRNA levels in the rVLM 24 and 48 hours after two applications of EA, separated by 72 hours. * P < 0.05 compared with the sham group.

Fig. 4

Concentration of met-enkephalin in the rVLM. Heights of bars represent the concentration measured from pooled brains of rats, with animal numbers shown above each bar.